| Budget Amount *help |
¥4,000,000 (Direct Cost: ¥4,000,000)
Fiscal Year 2000: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1999: ¥3,000,000 (Direct Cost: ¥3,000,000)
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| Research Abstract |
The results and research projects currently underway in our laboratory are as follows : (1) Methodology for analyses of glycosaminoglycans (GAGs). One of the most reliable methods, which are useful for determination of biological samples, is CE.We have already established determination methods for various GAGs involving CE with detection. NMR spectroscopy can provide quantitative and structural information of glycoconjugates such as glycosaminoglycans and proteoglycans. The proton NMR spectroscopic method has been established for determination of uronate residues in dermatan/heparan sulfates. (2) Preparation of oligosaccharides from a new glycosaminoglycan from the giant snail. A large amount of acharan sulfate was purified from the giant snail and the glycosaminoglycan was treated with heparin lyase II to obtain oligosaccharides. The oligosaccharides ranged 4-10 mers were purified by GPC and SAX HPLC methods, and each oligosaccharide was characterized by proton NMR.(3) The regulation
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of fibroblast growth factors by glycosaminoglycans. Fibroblast growth factors are important heparin binding, mitogenic proteins. The binding site in heparin and heparan sulfate for fibroblast growth factor-2 (basic fibroblast growth factor) has been described as rich in glucosamine-2-sulfate 1->4linked to iduronic acid-2-sulfate. The glucosamine residue in the heparin binding site is also 6-sulfated. A new glycosaminoglycan, acharan sulfate, has been chemically modified to prepare a polysaccharide, N-sulfoacharan sulfate, consisting of glucosamine-2-sulfate 1->4 linked to iduronic acid-2-sulfate. Acharan sulfate binds very weakly to fibroblast growth factor-2 while N-sulfoacharan sulfate binds with nearly the same affinity as heparin. Mitogenicity studies were performed using heparan sulfate-free cells stably transfected with fibroblast growth factor receptor-1. Acharan sulfate inhibits heparin's enhancement of fibroblast growth factor-2 mitogenic activity, without affecting cell viability, while N-sulfoacharan sulfate shows heparin-like activity but at a greatly reduced level. These results suggest additional mechanisms not requiring high affinity glycosaminoglycan binding to fibroblast growth factor-2 may be important in its mitogenic activity. Less
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